Composites material with suspended particles and method of using the same

ABSTRACT

An intermediate composite capable of transferring a biological or chemical material to be patterned on a surface. The intermediate composite includes a hydrogel, and particles suspended in the hydrogel, generating a particle-gel composite (composite), the composite is configured to absorb a biological or chemical material (agent), and further configured to deposit the agent when the composite is positioned proximate to a surface on which the agent is to be deposited.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present patent application is related to and claims the prioritybenefit of U.S. Non-Provisional patent application Ser. No. 14/824,577filed Aug. 12, 2015, and to U.S. Provisional Patent Application Ser. No.62/036,440, filed Aug. 12, 2014, the contents of each of which is herebyincorporated by reference in its entirety into this disclosure.

TECHNICAL FIELD

The present invention generally relates to micro-patterning andparticularly to micro-patterning a surface to functionalize the surface.

BACKGROUND

This section introduces aspects that may help facilitate a betterunderstanding of the disclosure. Accordingly, these statements are to beread in this light and are not to be understood as admissions about whatis or is not prior art.

Functionalizing solid flat surfaces or micro/nano sensor surfaces is awidely used procedure in many fields including biology, chemistry,medicine, and biomedical engineering. This procedure is usuallyperformed by pipetting liquids onto an area larger than the intendedarea (in order to ensure sufficient coverage). Patterning of surfaces inorder to result in gradients where the deposited pattern changes (i.e.,in amount or in a particular dimension) has been used in diverseapplications ranging from biosensing to cell culture systems. Severalmethods have been employed to create such gradients includingdiffusion-based microfluidics, laser desorption, and photochemistry.However, such methodologies often require complex equipment and harshtreatments which prevent the widespread use of gradient patterning.

Techniques such as micro-contact printing do exist but they require a“stamp” be prepared beforehand. The user cannot change the stamppattern. If the user decides that a new pattern is needed, the user hasto make a new stamp with the desired new pattern.

Therefore, there is an unmet need for a novel approach that canselectively and robustly functionalize a surface by depositingbiological or chemical traces on the surface.

SUMMARY

An intermediate composite capable of transferring a biological orchemical material to be patterned on a surface is disclosed. Theintermediate composite includes a hydrogel, and particles suspended inthe hydrogel, generating a particle-gel composite (composite), thecomposite is configured to absorb a biological or chemical material(agent), and further configured to deposit the agent when the compositeis positioned proximate to a surface on which the agent is to bedeposited.

A method of depositing biomolecules or chemical material on a surface isalso disclosed. The method includes suspending spores in a hydrogel,generating a hydrogel-spore composite (intermediate composite). Themethod also includes embedding biomolecule or chemical material (agent)into the intermediate composite thereby allowing absorption of the agentby the spores generating a composite (Composite). In addition, themethod includes micro-manipulating the Composite proximate to a surfaceresulting in deposition of the agent on the surface.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1(a) is a schematic illustration of a patterning procedureaccording to the present disclosure that is used to generateuser-defined surface concentrations.

FIG. 1(b) is a schematic illustration representing ability of negativelycharged spores to bind to a positively charged agent.

FIG. 1(c) is a schematic illustration of a micromanipulator depositingan agent on a surface according to the present disclosure.

FIG. 2 is a micrograph of a composite material being manipulated by amechanical micro-tweezers.

FIG. 3 is a micrograph showing bright field and fluorescent images of ahydrogel, and a hydrogel-spore composite when they are exposed to a dye.

FIG. 4 is a micrograph depicting a hydrogel-spore composite according tothe present disclosure, showing that the spores may tend to cluster nearthe surface of the hydrogel due to convective forces.

FIG. 5 is a plot of normalized fluorescent intensity vs. relativehumidity obtained from a humidity response experiment.

FIG. 6 is a micrograph showing volumetric changes in fluoresceinisothiocyanate (FITC)-loaded composite gel due to increase in humidity.

FIG. 7 is a schematic illustration of a model showing the mechanism bywhich spores can increase the concentration of chemical or biologicalagents inside the hydrogel.

DETAILED DESCRIPTION

For the purposes of promoting an understanding of the principles of thepresent disclosure, reference will now be made to the embodimentsillustrated in the drawings, and specific language will be used todescribe the same. It will nevertheless be understood that no limitationof the scope of this disclosure is thereby intended.

A novel approach that can selectively and robustly functionalize asurface by depositing biological or chemical traces on the surface isdisclosed. The approach includes a patterning strategy based upon aspore-hydrogel composite material, e.g., by suspending Bacillus subtilis(also referred to as B. subtilis) spores within a humidity sensitivehydrogel. The spore-hydrogel composite provides the capability to absorbchemicals and biological agents (generally referred to as agent),collectively (i.e., spore-hydrogel composite and agent) referred to asComposite, thereby generating a micro-environment that can bemanipulated by a micromanipulator in order to deposit the absorbed agentonto a surface. Deposition on to the surfaces can be carried out in avariety of user-defined patterns. For example, a user can functionalizea surface in a zig-zag, circular or other more complicated patterns. Onecan engineer the properties of the Composite by controlling the amountand the type of the spores introduced into the Composite. Moreover, bycontrolling the humidity in the environment, the user can also controlthe amount of agent that is released by the Composite. Hence, it ispossible to generate user-defined patterns with user-definedconcentrations. Controlling the humidity level is one example ofcontrolling the chemical release.

It should be understood that the approach described herein is notlimited to B. subtilis. Other spores can also be employed. Further, theapproach is not intended to be limited to spores. Materials other thanspores can be employed, such as spheres, cells, micro particles or nanoparticles that have organic or inorganic content.

Bacillus subtilis is a commonly found bacterium usually recovered fromwater, soil, air, and decomposing plant residue. The bacterium producesan endospore that allows it to endure extreme conditions of heat anddesiccation in the environment. B. subtilis produces a variety ofproteases and other enzymes that enable it to degrade a variety ofnatural substrates and contribute to nutrient cycling. However, undermost conditions the organism is not biologically active but exists inthe spore form. B. subtilis is considered a benign organism as it doesnot possess traits that cause disease.

A Hydrogels is a network of polymer chains that are generallyhydrophilic, sometimes found as a colloidal gel in which water is thedispersion medium. Hydrogels are highly absorbent (they can contain over90% water) natural or synthetic polymeric networks. Hydrogels alsopossess a degree of flexibility very similar to natural tissue, due totheir significant water content.

B. subtilis can swell and absorb water from humid environments inaddition to acting as a micro-carrier when brought into coupling with abiological/chemical agent. In the present disclosure, the ability of B.subtilis to act as such a carrier is demonstrated by loading B. subtilisspores that are embedded in a hydrogel forming the hydrogel-sporecomposite, altogether including the dye forming the Composite. Forexample, by saturating spores with a fluorescent dye and subjecting theComposite to various humidity levels, the concentration of dye releasedfrom the hydrogel surface can be controlled. In this disclosure it isdemonstrated therefore that the carrier (B. subtilis spores in thehydrogel-spore composite) can be used to engineer the concentration ofthe agent within the hydrogel-spore composite, and hence this approachcan be exploited as an additional parameter to control the agent releaseprocess. The fluorescent dye used here is an example of a material thatcan be used. As discussed above, it should be appreciated that the useof the dye is used inter alia to demonstrate the effectiveness of thenovel arrangement according the present disclosure, and that the dye canbe replaced with a variety of agents such as biomolecules includingproteins, DNA, RNA, small molecules, peptides, ions, salts, sugars,cells, and pathogens including viruses and bacteria.

FIG. 1(a) is a schematic illustration of a patterning procedure 100according to the present disclosure to generate user-defined surfaceconcentrations of an agent on a surface. In an exemplary embodiment, amicromanipulator 102 is used to handle Composite 104 and place theComposite 104 near a surface 106 according to a user-defined pattern.The Composite 104 as shown in FIGS. 1(a) and 1(c) includes a hydrogel120, spores 122 dispersed throughout the hydrogel 120 at a predefinedconcentration, and agent which includes agent molecules identified as124 in FIG. 1(c) and agent liquid identified as 128 in FIG. 1(c).

Referring back to FIG. 1(a), the concept of agent transfer is depicted.When the agent (including agent molecules 124 and agent liquid (see FIG.1(c)) has already been introduced to the hydrogel-spore compositegenerating the Composite 104, the agent molecules 124 (see FIG. 1(c))are carried by the spores 122 as well as by the hydrogel 120 which alsocarries the agent liquid 128. The agent including the agent molecules124 and agent liquid 128 is transferred to the surface 106 when theComposite 104 is brought proximate to the surface 106. The mechanism oftransfer is typically based on formation of a meniscus 126 (see FIG.1(c)) when the micromanipulator 102 brings the Composite 104 near thesurface 106 and when the humidity levels are at an appropriate level. Atlow humidity levels, the meniscus 126 does not form at the Composite 104and surface 106 interface leading to low levels of agent transfer (asshown in the upper panel of FIG. (1 a)). At mid-range humidity levels, aliquid meniscus forms allowing transfer of the agent (containing agentmolecules 124 and agent fluid 128) on to the surface 106 (middle panelof FIG. 1(a)). As the humidity increases further, the hydrogel 120absorbs more water and swells (bottom panel of FIG. 1(a)); thus,diluting the internal concentration of the agent (including the agentmolecules 124 and the agent liquid 128) inside the Composite 104, andthus decreasing the concentration of the agent that is transferred tothe surface 106.

To further improve the concentration of the agent within the Composite104, the agent molecules 124 can be electrically charged with a chargeopposite to the charge of the spores 122. FIG. 1(b) is a schematicillustration of the ability of negatively charged spores 122 to bind topositively charged agent molecules 124, thus increasing the internalconcentration of the agent molecules 124. The left panels of FIG. 1(b)depict the hydrogel with spores therein. The right panels of FIG. 1(b)depict the Composite 104 (i.e., hydrogel 120, spores 122, and the agent(i.e., the agent molecules 124 and the agent liquid 128)). In the toppanels of FIG. 1(b), the hydrogel 120 is shown in a mid-level humiditythereby allowing a high concentration of spores 122 and thereby a higherconcentration of the agent molecules 124. With the agent molecules 124positively charged, a high concentration of the agent molecules 124 canbe associated with the spores 122 and the hydrogel 120. In the bottompanels of FIG. 1(b), the hydrogel 120 is shown in a high-level humiditythereby resulting in a low concentration of spores 122 (bottom-leftpanel of FIG. 1(b)) and spores 122 and agent molecules 124 together(bottom-right panel of FIG. 1(b)). In other embodiments of thisdisclosure, the spores can be replaced with other suspended particles ofa material and with a certain charge that attracts dyes or otherchemicals of the opposite charge. Alternatively, spores can be replacedwith other particles that retain agents to be released by ways otherthan electrostatic interaction.

It should be appreciated that the direct use of spores to pattern onsurfaces proves difficult due to the brittle nature of spore aggregatesand the small size of spores (approximately 1 μm). The arrangementaccording to the present disclosure address this problem by suspendingthe spores in a stimuli-responsive hydrogel to improve the structuralintegrity of the spores while maintaining access to environmentalhumidity. In addition, the large size of the hydrogel construct (50-100μm) facilitates manual manipulation using micromanipulators, thusallowing easy, user-controlled patterning using the Composite 104.

FIG. 2 is a micrograph of the Composite 104 being manipulated by themechanical micromanipulator 102. The micromanipulator 102 allows forpatterning size selection through grasping different sizes of Composite104; in addition, the micromanipulator 102 gives the user precisespatiotemporal control over the patterning.

The hydrogel-spore composites were prepared by drying either 0.1 ml or0.2 ml of B. subtilis spores, 10⁷ CFU (MESA LABS) (the term CFU isgenerally used to mean colony forming units) and suspending theresultant spore powder into 0.9 ml of 25.9 mg acrylamide (SIGMA-ALDRICH)containing 75.2 μl methacrylic acid (SIGMA-ALDRICH, distilled to removeinhibitor), 12.2 mg N,N-methylenebisacrylamide (Polysciences Inc.), and75 μl N,N,N,N-tetramethylethylenediamine (SIGMA-ALDRICH) dissolved indeionized (DI) water. Gelation was induced by adding the solution into a0.1 ml aqueous solution of 0.35 M ammonium persulfate and subsequentlyvortexing to ensure uniformity in spore distribution. A negative chargecontrol was made by pipetting 0.5 μl poly(lactic-co-glycolic acid)(PLGA) microspheres (1% aqueous suspension of 1 μm microspheres,PHOSPHOREX INC.) into the solution prior to gelation in order to mimicthe charge and size of spores within the gel.

The binding and retention of avidin-fluorescein isothiocyanate (FITC)was verified by plating the hydrogel Composites on a glass slide,saturating the hydrogel with avidin-FITC (SIGMA-ALDRICH), and triplerinsing with running deionized water for 5 min. FIG. 3 is a micrographshowing bright field and fluorescent images of hydrogel-only control,and loaded hydrogel-spore composites (i.e., Composites). In FIG. 3, thetop row shows bright field and fluorescent micrographs of hydrogelcontrol (left panel without any agent introduced, middle panel with dyeagent introduced, and right panel representing the status after a washwith H₂O) while the bottom two rows show hydrogel-spore composites. Theimages in the left column correspond to a stage before the addition ofavidin-FITC, the images in the middle column correspond to a stage aftersuch addition, and the right column represents images after a 1-minutewash with water. These images demonstrate the binding of dye is at leastpartially charge dependent (difference between the middle row,particularly the right panel, and the bottom row, particularly the rightpanel). The gel constructs can be visualized as the dark area on theright-hand side of the bright field images.

FIG. 3 shows that the hydrogel-spore composite retains dye near the gelsurface while a hydrogel-only control shows no significant retention ofdye. When the hydrogel-spore composite was examined under bright fieldmicroscopy, spores were seen at very high concentrations near thesurface of the gel. The concentration of spores decayed quickly from thesurface to the hydrogel center. Since the hydrogel in this controlexperiment was synthesized directly on the surface of the glass slide,it is believed that the spores were pushed towards the hydrogel surfaceby convective forces and were subsequently set in place when gelationoccurred. The high concentration of spores on the outer edge of thehydrogel-spore composite explains why fluorescence was seen only in thevicinity of the surface after rinsing. The water removed all dye fromthe hydrogel, except the dye protected by the spore coat. Anothercontrol was prepared by saturating the hydrogel-spore composite with a 1M magnesium chloride solution in order to mask the negative surfacecharge of the spores. After the rinse, the magnesium control showedessentially no retention of dye, suggesting that the spore surfacecharge aids in the retention of the dye.

FIG. 4 is a micrograph depicting the hydrogel-spore composite, showingthat the spores tend to cluster near the surface of the gel due toconvective forces present during synthesis.

The humidity response of hydrogels with different concentrations ofspores was characterized. Experiments were conducted with hydrogelsdevoid of spores (hydrogel control) and those that had negativelycharged PLGA microspheres instead of spores (charge control). Ahumidifier and a hygrometer were used to produce and measure thehumidity in a chamber. First, hydrogel samples were saturated withavidin-FITC and allowed to dry. Small aliquots of each gel(approximately 50 μm in diameter) were grasped using a mechanicalmicromanipulator. The gel was placed in the humidity chamber for 2 minprior to patterning to acclimatize to the environment. However, theglass slide remained outside the chamber until immediately prior topatterning in order to avoid condensation on the surface. The gel waslowered until a meniscus formed on the glass slide, patterning a smallcircular spot on the surface. The gel sample was re-soaked fully foreach humidity point to eliminate the effects of dye depletion from thisexperiment. The slide fluorescence profiles were immediately imaged tominimize photobleaching and were imported into a MATLAB algorithm foranalysis. The intensity of each image was normalized to that of thebackground intensity of the glass slide and was averaged over thepatterned area. This procedure was repeated for five different relativehumidity levels, and three data points were obtained at each humiditylevel.

FIG. 5 is a plot of normalized fluorescent intensity vs. relativehumidity obtained from the humidity response experiment described abovein summary form. In FIG. 5, the error bars shown represent standarddeviation of 3 experiments. FIG. 5 shows a large increase in intensityas the hydrogel is hydrated by increased relative humidity followed by agradual decline as the concentration of the spores and thereby theconcentration of the dye is decreased. Referring to FIG. 5, allexperiments showed a rapid increase in fluorescent intensity followed bya gradual decline as the relative humidity was increased from 35% to95%. The 0.2 ml hydrogel-spore composite showed the greatest increase influorescent intensity with 32% increase over that of the hydrogelcontrol. Using unpaired, 2-tailed, heteroscedastic t-tests (P<0.1),significant difference was seen between the Composites and variouscontrols. The 0.1 ml hydrogel-spore composite was statisticallydifferent from the charge control at 55% and 70% relative humidity andfrom the hydrogel control at 85% relative humidity. The 0.2 mlhydrogel-spore composite was statistically different from both controlsfor all relative humidity values except 35%. At 35% relative humidity,the dry gel-spore composite did not have enough moisture to form ameniscus to transfer dye to the surface, leading to low initialfluorescent intensity. However, at higher humidity levels, the hydrogelbegan to absorb ambient humidity leading to the formation of a meniscusand subsequent deposition of the fluorescent dye. As the humiditycontinued to increase, the gel significantly swelled with water,increasing in size and slowly diluting the concentration of dye withinthe gel, hence resulting in a decreased deposition of the fluorescentdye. FIG. 6 is a micrograph showing volumetric changes in theFITC-loaded Composite due to increase in humidity.

The level of fluorescent intensity deposited on the surface using theComposite is directly attributed to the ability of the Composite to bindhigher concentrations of dye compared to the hydrogel and the chargecontrol gel, thus creating a steeper gradient as the humidity is varied.The binding and retention experiment showed that charge plays a role inthe ability of spores to bind higher levels of dye than hydrogel alone;however, the statistical difference from the charge control of thehumidity response experiment suggests that the mechanism involves morethan simple electrostatic interaction. Thus, there is a positivecorrelation between the peak concentration of patterned dye and theconcentration of spores present in the Composite.

It should be noted that the amount of negatively charged surface areawithin the gel affects the rate at which surface concentration decreasesas humidity increases. In the hydrogel control, as the humidityincreases, the internal concentration decreases due to the increasedvolume of water. In the Composites, as the humidity increases, theinternal concentration decreases as well. However, the dye bound to thecharged surface has an increased diffusive flux due to the decreasedconcentration within the surrounding gel. This increases the diffusionfrom the spore to the surrounding gel, which attenuates theconcentration lowering effects of increased humidity. FIG. 7 is aschematic illustration of a model showing the mechanism by which sporescan attenuate the concentration lowering effects over mid-to-highhumidity range compared to a hydrogel control.

As described above, a novel method of patterning biomolecules on asurface has been developed. The method includes suspending spores in ahydrogel forming a Composite, embedding a set of biomolecules desired tobe deposited onto a surface into the Composite, and micro-manipulatingthe Composite on the surface resulting in a deposition of thebiomolecules on the surface. In one embodiment, the amount ofbiomolecules deposited is dependent on the humidity prevailing in theComposite. The humidity levels in the Composite can be controlled asdescribed above. In one embodiment, for example, the spores can beBacillus Subtilis. In other embodiments, spores other than Bacillussubtilis, such as Bacillis anthracis or other bacteria can be used tomodify the properties of the hydrogel substrate. The spores or bacteriacan be chosen based on the electrostatic, mechanical, optical,topographical, geometrical, material or chemical properties that mayaffect the type or the amount of the target dye or material that is tobe released. In some embodiments of this disclosure, the use of sporescan be totally replaced with the use of non-biological particles such asspheres, micro particles or nano particles that have organic orinorganic content including but not limited to magnetic particles,polystyrene particles, quantum dots that may attract and retain agents(e.g., biomolecules or chemicals to be released) by electrostatic,magnetic, steric, hydrophilic, hydrophobic, mechanical or chemicalmeans. Further, in some embodiments, the material to be released to thesurface can be biomolecules such as proteins, DNA, RNA, peptides,peptide nucleic acids, or small molecules, sugars, salts or ions. Itshould be further recognized, that in embodiments of this disclosure,the surface that receives the deposited material can be glass, silicon,silicon dioxide, indium tin oxide, polymeric, plastic, or metallic(meaning metal or alloy), including but not limited to gold, platinum,and other metals. The surface can be flat or irregular since theapproach described in this disclosure allows depositing materials ontonon-flat surfaces as well. For example, in one embodiment, the systemcan be used to deposit materials to the side of an object or selectivelyon the oblique planes of a prism like object. Further, it is envisionedthat the surface can be that of a sensor or actuator or another devicethat needs the presence of certain chemicals or molecules or materialson it for operability or enhanced performance. Furthermore, whilechanges in humidity were accomplished by placing the spore-gel in ahumidity chamber, selective humidity can be implemented on the surfaceby selectively applying the appropriate amount of vapor directly at thesurface. While humidity was used as a stimulus in one of the embodimentsdescribed above to deposit and control the material, the stimulus thatcontrols the material release need not be limited to humidity. Forexample, the release can be triggered or controlled by temperature,electric fields, magnetic fields, mechanical forces or by introductionof or exposure to other chemicals. It should be noted that such stimulican be combined. For example, the stimuli of temperature and humiditycan be used in combination to control the amount of material releasedfrom the Composite. In the approach described in this disclosure, thesubstrate material of the Composite may be different from a hydrogel andmay include other gels, polymers or solids that can be manipulated by apair of tweezers or similar manipulators. Also while a micromanipulatorwas discussed in the present disclosure for the purpose of manipulatingthe position of the Composite on to the surface, other devices andarrangements may be used to bring the Composite at a larger scale on tothe surface.

While in this disclosure embodiments containing one Composite isdescribed, it should be appreciated that multiple Composites can beemployed in a patterning process of the type described in thisdisclosure. Further, the multiple Composites can contain varyingchemistries or physical characteristics for the suspended particles.Furthermore, in utilizing multiple Composites and/or multiplechemistries or physical characteristics for the suspended particles,more than one stimulus can be utilized advantageously. The stimulus orstimuli can be chosen based on the effect of the stimulus or stimuli onthe Composites and/or the suspended particles. Thus in a singlepatterning exercise, it is possible to combine a variety of Composites,a variety of suspended particles and a variety of stimuli. Thesepossibilities can be exploited to control the rate of deposition and/orconcentration gradients of the deposited materials.

It should be appreciated that the deposited agent, according to thepresent disclosure, can be deposited in a manner in which it generates adiffraction grating pattern. The agent can provide an affinity forbinding to biological entities of particular variety, e.g., cellsafflicted with a particular disease. The agent can then bind to thosebiological entities when a fluid containing those biological entities ispassed over the deposited agent thereby generating a binding affinity tothe agent. The diffraction grating pattern can then be used to quicklyidentify the bound biological entities using techniques known to aperson having ordinary skill in the art.

It should be appreciated that the arrangement described herein fordepositing the agent can be provided in an array. In other words, anarray of micromanipulators can be grouped together to deposit variouspatterns of the agent on to the surface according to a user-definedpattern. In this embodiment, a larger pattern can be deposited in ashorter amount of time.

It should also be appreciated that the surface deposition of the agentcan be accomplished on a combination of surface orientations, e.g.,horizontal, vertical, angled, and any combination thereof.

Those having ordinary skill in the art will recognize that numerousmodifications can be made to the specific implementations describedabove. The implementations should not be limited to the particularlimitations described. Other implementations may be possible.

1. An intermediate composite capable of transferring a biological orchemical material to be patterned on a surface, comprising: a hydrogel;and particles suspended in the hydrogel, generating a first composite,the composite is configured to absorb a biological or chemical material(agent) thereby generating a second composite, and further configured todeposit the agent when the second composite is positioned proximate to asurface on which the agent is to be deposited.
 2. The intermediatecomposite material of claim 1, wherein the suspended particles arespores.
 3. The intermediate composite material of claim 2, wherein thesuspended particles are Bacillus subtilis spores.
 4. The intermediatecomposite material of claim 1, wherein the concentration of theparticles in the hydrogel is between about 0 spores/mL and about 1×10¹²spores/mL.
 5. The intermediate composite material of claim 4, whereinthe concentration of the particles in the hydrogel is between about 10⁷spores/mL and about 2×10⁷ spores/mL.
 6. The intermediate compositematerial of claim 1, wherein each of the first composite or the secondcomposite is sensitive to humidity.
 7. The intermediate compositematerial of claim 6, wherein the humidity at each of the first compositeor the second composite can be varied from about 3.5% to about 95%. 8.The intermediate composite material of claim 1, the agent is configuredto be deposited on any combination of vertical and horizontal surfaces.9. The intermediate composite material of claim 1, the agent isconfigured to be deposited according to a user-defined pattern.
 10. Theintermediate composite material of claim 9, wherein the agent isconfigured to be deposited generating a concentration gradient of theagent on the surface.
 11. A composite capable of transferring abiological or chemical material to be patterned on a surface,comprising: a hydrogel; and particles having a first charge suspended inthe hydrogel, generating a first composite, the first composite isconfigured to absorb a biological or chemical material (agent) having asecond charge thereby generating a second composite, and furtherconfigured to deposit the agent when the second composite is positionedproximate to a surface on which the agent is to be deposited.
 12. Thecomposite material of claim 11, wherein the suspended particles arespores.
 13. The composite material of claim 12, wherein the suspendedparticles are Bacillus subtilis spores.
 14. The composite material ofclaim 11, wherein the concentration of the particles in the hydrogel isbetween about 0 spores/mL and about 1×10¹² spores/mL.
 15. The compositematerial of claim 14, wherein the concentration of the particles in thehydrogel is between about 10⁷ spores/mL and about 2×10⁷ spores/mL. 16.The composite material of claim 11, wherein each of the first compositeor the second composite is sensitive to humidity.
 17. The compositematerial of claim 16, wherein the humidity at each of the firstcomposite or the second composite can be varied from about 3.5% to about95%.
 18. The composite material of claim 11, the rate of agentdeposition is dependent on i) humidity, ii) the first and second charge,or iii) concentration of the particles in the hydrogel.
 19. Thecomposite material of claim 18, wherein the first charge-second chargeis one of negative-positive, positive-negative, positive-positive,negative-negative, neutral-positive, neutral-negative, negative-neutral,or neutral-neutral.
 20. The composite material of claim 19, the rate ofagent deposition increases by about 20 percent when humidity increasesfrom about 35% to about 55% when the first charge-second charge isnegative-positive.